The present invention relates to a vaccine that comprises liposomes adsorbed to aluminum hydroxide, the liposomes comprising lipid A and a molecule ("malarial antigen") that is capable of inducing an immune response against a malarial parasite in man or in an animal species. The present invention also pertains to a method of inducing an immune response in a mammal against a malarial parasite by injection of the above-described vaccine.
In the normal course of malarial infection, the parasitic malarial organism, in the form of a sporozoite, is transmitted to an animal host by the bite of a mosquito. Shortly thereafter, the sporozoite invades the host's liver where it replicates and produces numerous merozoites. These merozoites then invade the erythrocytes of the host.
In the development of a vaccine for protection of man and animals against malaria, therefore, it is desirable to produce a vaccine that can induce immunity against one of the life forms of the malarial parasite. For example, a vaccine can be developed against the sporozoites and thereby prevent invasion of the liver tissue and replication of the parasite. But in order for the induced immune response to be effective, the anti-sporozoite antibody titer must be sufficiently high (1) to maintain protective activity over a long period of time, preferably more than a year, and (2) to block sporozoite invasion of the liver within the short period of time after infection and before invasion of the liver.
Sporozoites are known to possess an antigenic surface protein known as a circumsporozoite ("CS") protein. The gene encoding the CS protein from Plasmodium falciparum has been cloned, and the amino acid sequence of this CS protein has been determined. It has been found that the middle portion of the CS protein comprises a region that has thirty-seven Asn-Ala-Asn-Pro tetrapeptides interspersed with four Asn-Val-Asp-Pro tetrapeptides. Peptides comprising various multiples of at least one of these two tetrapeptides are herein collectively referred to as "CS peptides."
Young et al, Science 228: 958-962 (1985), found that the highest antibody titer that could be induced in mice against the malarial parasite occurred when these animals were injected with CS peptides in admixture with complete Freund's adjuvant ("CFA"). When the mice were immunized with CS peptides adsorbed to aluminum hydroxide, without CFA, an intermediate level of antibody titer was obtained. Injection of one CS peptide, R16tet.sub.32, alone into mice induced very little immune response. It appears, therefore, that CRA is capable of enhancing the immune response of mice to a non-immunogenic CS peptide.
In the present description, "R16tet.sub.32 " denotes a synthetic peptide that has (1) fifteen Asn-Ala-Asn-Pro tetrapeptide repeats and one Asn-Val-Asp-Pro tetrapeptide and (2) thirty-two amino acids derived from a tetracycline resistance gene. In like fashion, "R32tet.sub.32 " and "R48tet.sub.32 " denote peptides that are similar to R16tet.sub.32 but that have two and three copies, respectively, of the sixteen tetrapeptides of R16tet.sub.32.
For purposes of vaccination in human, use of CFA is not desirable because of its severe side effects. Therefore, alternative adjuvants for stimulating the immunogenicity of antigens for use in man are sought. Liposomes have been used as an alternative adjuvant in animal systems. The ability of different liposomes to enhance immune response, however, has been variable.
For example, Allison and Gregoriadis, Nature 252: 252 (1974), reported that the use of negatively charged or neutral liposomes effectively induced a higher immune response to diphtheria toxoid ("DT") in mice, but that use of positively charged liposomes led to a weaker immune response than was obtained with the use of DT alone. In other studies in which other antigens were used, positively charged and negatively charged liposomes were found to be equally effective as adjuvants. See Heath et al., Biochem. Soc. Trans. 4: 129-133 (1976), and van Rooijen and van Nieuwmegen, Immunol. Commun. 9: 243-256 (1980). Accordingly it appears that whether liposomes can act as adjuvant varies from antigen to antigen.
Alving et al., Vaccine 4: 166-172 (1986) used liposomes comprising lipid A as adjuvants in inducing immunity in rabbits to cholera toxin ("CT") and to a synthetic CS peptide consisting of four Asn-Pro-Asn-Ala tetrapeptides conjugated to BSA. The authors found that the immune response to CT or to the synthetic malaria peptide was markedly enhanced by use of liposomes and lipid A, compared to similar liposomes lacking lipid A.
In contrast, Gerlier et al., J. Immunol. 131: 485-490 (1983), found in rats that liposomes comprising the same lipid A as used by Alving and his coworkers did not stimulate a greater immune response to antigens derived from cells infected with Gross virus. Hence, it again appears that the ability of liposomes with lipid A alone to serve effectively as adjuvants varies depending on the antigen and, perhaps, on the animal system used.